Centrifuge system for separating cells in suspension

ABSTRACT

A method and apparatus for separation of cells from suspension using single use components that include a flexible membrane mounted on a rigid frame that is supported within a multiple use rigid centrifuge bowl. Such single use components include a rotatable core which is rotated at a sufficiently high angular velocity to separate cell concentrate and centrate. Cell centrate and/or cell concentrate is removed from the interior of the core by a centripetal pump having a pair of curved volute passages. The volute passages are curved toward the direction of rotation of the core.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims benefit pursuant to 35 U.S.C. § 119(e) ofProvisional Application Ser. No. 62/163,433 filed May 19, 2015.

This application is a continuation-in-part of application Ser. No.14/698,995 filed Apr. 29, 2015, which application is a continuation ofapplication Ser. No. 13/684,051 filed Nov. 21, 2012 which applicationclaims benefit under 35 U.S.C. § 119(e) of Provisional Application61/562,438 filed Nov. 21, 2011.

Application Ser. No. 13/684,051 is a continuation-in-part of U.S.application Ser. No. 12/676,273 filed Mar. 3, 2010 which claims benefitunder 35 U.S.C. § 371 of PCT/US2009/002464 filed Apr. 21, 2009 whichclaims benefit under 35 U.S.C. § 119(e) of Provisional Application61/125,033 filed Apr. 22, 2008.

The disclosure of each prior application is incorporated herein byreference in its entirety.

TECHNICAL FIELD

This disclosure relates to centrifugal processing of materials.Exemplary embodiments relate to devices for separating cells insuspension through centrifugal processing.

BACKGROUND

Devices and methods for centrifugal separation of cells in suspensionare useful in many technological environments. Such systems may benefitfrom improvements.

SUMMARY

The exemplary embodiments described herein include apparatus and methodsfor centrifugal separation of cells in large-scale cell culture with ahigh cell concentration using pre-sterilized, single-use fluid pathcomponents. The exemplary centrifuges discussed herein are solid wallcentrifuges that use pre-sterilized, single-use components, and arecapable of processing cell suspensions, with high cell concentrations,at flow rates in the range of about 2 to about 40 liters per minute,preferably about 5 to about 20 liters per minute.

The exemplary embodiments use rotationally fixed feed and dischargecomponents. Single use components include a flexible membrane mounted ona rigid frame including a core with an enlarged diameter. The single usecomponents further include at least one centripetal pump. The single usestructures are supported within a multiple use rigid bowl having aninternal truncated cone shape. These structures permit the exemplarysystems to maintain a sufficiently high angular velocity to create asettling velocity suited to efficiently processing highly concentratedcell culture streams. Features which minimize feed turbidity, and otherswhich permit the continuous or semi-continuous discharge of cellconcentrate, increase the overall production rate over the rate whichcan be achieved. Exemplary structures and methods provide for effectiveoperation and reduce risks of contamination.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a schematic view of an exemplary embodiment of a centrifugesystem including single use and multiple use components.

FIG. 2 is a close-up view of the upper flange area of the centrifuge ofFIG. 1, which shows a method of sealing the flexible chamber material tothe surface of the flange.

FIG. 3 is an isometric cutaway view of the core and upper flanges of thesingle use component of the embodiment the centrifuge system of FIG. 1.

FIG. 4 is a schematic view of the embodiment illustrated in FIG. 1, inwhich the pump chamber of the centrifuge system includes acceleratorfins.

FIG. 5 is an isometric view of the top of the pump chamber of theembodiment of the centrifuge system illustrated in FIG. 4.

FIG. 6 is an isometric cutaway view of the core, upper flanges and lowerflanges, of a single use centrifuge system with an enlarged corediameter (to create a shallow pool centrifuge), and a feed accelerator.

FIG. 7 is an isometric view of the feed accelerator of FIG. 6.

FIG. 8 is an isometric cutaway view of the core and upper flanges of asingle use centrifuge system with a standard core diameter, and a feedaccelerator with curved vanes and an elliptical bowl.

FIG. 9 is an isometric view of the feed accelerator of FIG. 8.

FIG. 10 is a schematic view of a portion of a continuous concentratedischarge centrifuge system.

FIG. 11 is a schematic view of a portion of a second embodiment whichincludes a continuous concentrate discharge centrifuge system.

FIG. 12 is a schematic view of a continuous concentrate dischargecentrifuge system with diluent injection.

FIG. 13 is schematic view of a portion of a third embodiment of acontinuous concentrate discharge system, with a throttle mechanism forthe centripetal pumps.

FIG. 14 is an isometric cutaway view of the core and upper flanges of asingle use centrifuge system with a standard core diameter, and a feedaccelerator with straight vanes.

FIG. 15 is an isometric view of the feed accelerator of FIG. 14.

FIG. 16 is an isometric cutaway view of an alternative continuousconcentrate discharge centrifuge system.

FIG. 17 is an isometric exploded view of an alternative centripetalpump.

FIG. 18 is an isometric view of a plate of the alternative centripetalpump including the volute passages therein.

FIG. 19 is a schematic view of a centrifuge system which operates toassure that positive pressure is maintained in the centrifuge corecavity.

FIG. 20 is a schematic view showing simplified exemplary logic flowexecuted by at least one control circuit of the system shown in FIG. 19.

DETAILED DESCRIPTION

In the field of cell culture as applied to bio-pharmaceutical processesthere exists a need to separate cells from fluid media such as fluid inwhich cells are grown. The desired product from the cell culture may bea molecular species that the cell excretes into the media, a molecularspecies that remains within the cell, or it may be the cell itself. Atproduction scale, the initial stages of cell culture process typicallytake place in bioreactors, which may be operated in either batch orcontinuous mode. Variations such as repeated batch processes may bepracticed as well. The desired product must eventually be separated fromother process components prior to final purification and productformulation. Cell harvest is a general term applied to these cellseparations from other process components. Clarification is a termdenoting cell separations in which a cell-free supernatant (or centrate)is the objective. Cell recovery is a term often applied to separationswherein a cell concentrate is the objective. The exemplary embodimentsherein are directed to cell harvest separations in large-scale cellculture systems.

Methods for cell harvest separations include batch, intermittent,continuous and semi-continuous centrifugation, tangential flowfiltration (TFF) and depth filtration. Historically, centrifuges forcell harvest of large volumes of cell culture at production scale arecomplex multiple use systems that require clean-in-place (CIP) andsteam-in-place (SIP) technology to provide an aseptic environment toprevent contamination by microorganisms. At lab scale and for continuouscell harvest processes, smaller systems may be used. The UniFugecentrifuge system, manufactured by Pneumatic Scale Corporation,described in published application US 2010/0167388, the entiredisclosure of which is incorporated herein by reference, successfullyprocesses culture batches for cell harvest in the range of 3-30liters/minute in quantities of up to about 2000 liters usingintermittent processing. Also incorporated herein in their entirety areU.S. patent application Ser. No. 14/698,995 filed Apr. 29, 2015; andSer. No. 13/684,051 filed Nov. 21, 2012, which are also owned byPneumatic Scale Corporation, the assignee of the present application.Intermittent processing generally requires periodically stopping bothrotation of the centrifuge bowl and the feed flow in order to dischargeconcentrate. This approach usually works well with lower concentration,high viability cultures, in which large batches can be processed, andthe cell concentrate discharged relatively quickly and completely.

There is sometimes a requirement to harvest cells from highlyconcentrated and/or low viability cell cultures, which contain a highconcentration of cells and cell debris in the material feed, which aresometimes referred to as “high turbidity feeds.” Such high turbidityfeeds can slow down the processing rate in some centrifugal separationsystems, because:

-   -   1. a slower feed flow rate is required to provide increased        residence time in the centrifuge in order to separate small cell        debris particles, and    -   2. the higher concentrations of both cells and cell debris may        result in the bowl filling rapidly with cell concentrate, which        requires the bowl to be stopped to discharge concentrate.        These combined factors may result in a reduced net throughput        rate, and unacceptably long cell harvest processing times. In        addition to the increased costs which may be associated with a        longer processing time, increased time in the centrifuge may        also result in a higher degree of product contamination and loss        in the harvesting low viability cell cultures.

A high concentration of cell and cell debris in a material feed may alsoresult in a cell concentrate with a very high viscosity. This may makeit more difficult to completely discharge the cell concentrate from thecentrifuge, even with a prolonged discharge cycle. In some cases, anadditional buffer rinse cycle may be added to obtain a sufficientlycomplete discharge of concentrate. The need to make either or both ofthese adjustments to the discharge cycle further increases theprocessing time, which can make the challenges of processing a largevolume of cell culture more complex and costly.

Scaling up the size of systems, by increasing the bowl size to increasethe length of the feeding portion of the intermittent processing cycleis sometimes not practical because it also results in a proportionatelylonger discharge cycle for the cell concentrate. Another limitation thatmay preclude simple geometric scale-up is variation in scaling of thepertinent fluid dynamic factors. The maximum processing rate of anycentrifuge depends on the settling velocity of the particles beingseparated. The settling velocity is given by a modification of Stokes'law defined by Equation 1:

$\begin{matrix}{v = \frac{\Delta \; {p \cdot r \cdot d^{2} \cdot \omega^{2}}}{18 \cdot \mu}} & {{Equation}\mspace{14mu} 1}\end{matrix}$

where v=settling velocity, Δρ is solid-liquid density difference, d isparticle diameter, r is radial position of the particle, ω is angularvelocity, and μ is liquid viscosity. With respect to scale-up geometry,changing the radius of the bowl changes the maximum radial position rthat particles can occupy. Therefore, if the other parameters inEquation 1 are held constant, an increase in bowl radius leads to anincrease in average settling velocity and a gain in throughput for agiven separation efficiency. However, as the radius increases it becomesmore difficult to maintain the angular velocity of the bowl because ofthe increased material strength that may be required, and otherengineering limitations. If a decrease in angular velocity is largerthan the square root of the proportional increase in radius, then theaverage settling velocity and the gain in throughput (which isproportional to radius) both decline.

One of the engineering limitations that must be considered is that theangular velocity needed to rotate the larger bowl may not be practicalto achieve because of the more massive and costly centrifuge driveplatform that would be needed.

In addition if the angular velocity is held constant as the radiusincreases, the forces urging the cells toward the walls of thecentrifuge also increase. When the bowl is rotated at sufficiently highangular velocity to create the desired processing efficiency, the wallsof the container and the cells which accumulate there, experience addedstress. As to the cells, this can cause cell damage by packing the cellsto excessively high concentrations. Cell damage is a drawback inapplications wherein cell viability needs to be maintained and can leadto contamination of products that are present in solution in thecentrate. The higher viscosity resulting from excessively high cellconcentrations is also sometimes a drawback for complete discharge ofthe cell concentrate.

Exemplary embodiments include apparatus and methods for continuous orsemi-continuous centrifugal separation of low viability cell suspensioncultures containing a high concentration of cells and cell debris, at arate suitable for processing large volumes of cell suspensions on acommercial scale. Some exemplary centrifuges are of pre-sterilized,single-use designs and are capable of processing such cell suspensionsat flow rates exceeding 20 liters per minute. This flow capacity enablestotal run times in the range of 2 to 3 hours for a 2000 literbioreactor. Exemplary embodiments of the single-use centrifuge systemsare capable of processing about 300 to 2,000 liters of fluid whileoperating at a rate of about 2 to 40 liters per minute.

FIG. 1 discloses a single use centrifuge structure 1000. The centrifugestructure 1000 includes a core structure 1500 (best shown in FIG. 3)comprising a core 1510, upper flanges 1300, lower flanges 1200, and aflexible liner 1100 sealed to both an upper flange 1300 and a lowerflange 1200. The centrifuge structure 1000 also includes a centripetalpump 1400, comprising a pair of stationary paring disks 1410 in arotating pump chamber 1420, and a rotating mechanical seal 1700.

Centrifuge structure 1000 also includes a feed/discharge assembly 2000.The assembly 2000 comprises a plurality of concentric tubes about therotational axis 1525 (labeled in FIG. 12) of the centrifuge 1000. Theinnermost portion of feed/discharge assembly 2000 includes a feed tube2100. A plurality of additional tubes concentrically surround the feedtube 2100, and may include tubes or fluid pathways to permit centratedischarge 2200, concentrate discharge 2500 (see, for example, FIG. 12),or diluent feed 5000 (see, for example, FIG. 12). Each portion of thefeed/discharge connection may be in fluid connection with a portion ofthe interior of the centrifuge 1000, and a collection or feed chamber(not shown) via appropriate fluid connections, and may include furthertubes which are in fluid connection with the concentric tubes to removeor add the centrate, concentrate, or diluents from or to the system.

The upper and lower flanges 1300, 1200, as illustrated in FIG. 1,comprise conical bowls, axially aligned with and concave toward the core1510. Core 1510 comprises a generally cylindrical body with a hollowcylindrical center large enough to accept feed tube 2100 having an axis1525 (labeled in FIG. 12). The upper flange 1300, the core 1510 and thelower flange 1200 may be a unitary structure to provide a strongersupport structure for flexible liner 1100 which is alternativelyreferred to herein as a membrane. In other embodiments, the corestructure 1500 may be formed from a plurality of component parts. Infurther embodiments, the core 1510 and upper flange 1300 may comprise asingle component, with a lower flange 1200 comprising a separatecomponent, or the core 1510 and lower flange 1200 may comprise a singlecomponent with the upper flange 1300 comprising a separate component.

An embodiment of a unitary core 1510 and upper flange 1300 isillustrated in FIG. 3. This unitary component would be joined to lowerflange 1200 to create the internal supporting structure 1500 of thesingle use components of centrifuge 1000. This structure anchors theflexible liner 1100 around a fixed internal rigid or semi-rigid supportstructure 1500 at both the top and bottom. When the centrifuge system isin use, the flexible liner 1100 is also supported externally by thewalls and cover of the multiple use structure 3000.

The exemplary separation chamber 1550 is an open chamber which isroughly cylindrical in shape, bounded roughly by the exterior surface1515 of the core 1510 and the flexible liner 1100, and by the uppersurface 1210 of the lower flanges 1200 and the lower surface 1310 of theupper flanges 1300. The separation chamber 1550 is in fluid connectionwith the feed tube 2100 via holes 1530 extending from the central cavity1520 of the core 1510 to the exterior surface 1515 of the core 1510. Theseparation chamber 1550 is also in fluid connection with the pumpchamber 1420 via similar holes 1540 through the core structure 1500. Inthis example, holes 1540 angle upward, toward the pump chamber 1420,opening into the separation chamber 1550 just below the junction betweenthe core 1510 and upper flanges 1300. As shown in FIG. 12, holes 1420 or4420 may enter pump chambers at an angle other than upward, includinghorizontally or at a downward angle. In addition, in some embodimentsholes 1420, 4420 may be replaced by slits, or gaps between acceleratorfins.

FIG. 1 also shows a feed/discharge assembly 2000 which includes a feedtube carrier 2300, through which feed tube 2100 extends into theposition shown in FIG. 3, close to the bottom of centrifuge structure1000. In this position the feed tube 2100 can perform both feed anddischarge functions without being moved. Shearing forces during thefeeding process may be minimized by careful design of the gap betweenthe nozzle 2110 of the feed tube 2100 and the upper surface 1210 of thelower flanges 1200, the diameter of the nozzle 2110 of the feed tube2100, and the angular velocity of the centrifuge. U.S. Pat. No.6,616,590, the disclosure of which is fully incorporated herein byreference, describes how to select appropriate relationships to minimizethe shearing forces. Other suitable feed tube designs which minimize theshearing forces associated with feeding a liquid cell culture into arotating centrifuge which are known to those skilled in the art may alsobe used.

FIG. 1 further includes a centripetal pump 1400 for discharging centratethrough a centrate discharge path 2200. In the embodiment shown in FIG.1, the centrate pump 1400 is located above the upper flange 1300 in apump chamber 1420. Pump chamber 1420 is a chamber defined by the uppersurface 1505 of the core 1510 and the inner surfaces 1605, 1620 of acentrifuge cover 1600. The centrifuge cover 1600 may include cylindricalwalls 1640 and a mating cap portion 1610 shaped like a generallycircular disk (shown in FIG. 5). The centrifuge cover 1600 may be formedas a unitary body, or from separate components.

As discussed in more detail below, in other embodiments, the shape andposition of the centrate pump chamber 1420 may vary. Chamber 1420 willgenerally be an axially symmetric chamber near the upper end of the corestructure 1500 which is in fluid connection with the separation chamber1550 via holes or slits 1530 which extend from adjacent the exterior ofthe core 1515 into the centrate pump chamber 1420. In some embodiments,as shown most clearly in FIGS. 11 and 12, centrate pump chamber 1420 maybe located in a recess within chamber 1550.

Exemplary centrate pump 1400 comprises a pair of paring disks 1410.Paring disks 1410 are two thin circular disks (plates), which areaxially aligned with the axis 1525 of core structure 1500. In theembodiment illustrated in FIGS. 1-5, paring disks 1410 are heldstationary relative to the centrifuge structure 1000, and are separatedfrom each other by a fixed gap 1415 (labeled in FIG. 10). The gap 1415between the paring disks 1410 forms part of a fluid connection forremoving centrate from the centrifuge 1000, which permits centrate toflow between the paring disks 1410 into a hollow cylindrical centratedischarge path 2200 surrounding the feed tube carrier 2300, terminatingin centrate outlet 2400.

The single use centrifuge structure 1000 is contained within a multipleuse centrifuge structure 3000. The structure 3000 comprises a bowl 3100and a cover 3200. The walls of the centrifuge bowl 3100 support theflexible liner 1100 of centrifuge structure 1000 during rotation of thecentrifuge 1000. In order to do so, the external structure of the singleuse structure 1000 and the internal structure of the multiple usestructure conform to each other. Similarly, the upper surface of upperflanges 1200, the exterior of an upper portion of core 1510, and a lowerportion of the walls 1640 of the centrifuge cover 1600 conform to theinner surface of the multiple use bowl cover 3200, which is also adaptedto provide support during rotation. Features of the multiple use bowl3100 and bowl cover 3200, discussed in more detail below, are designedto ensure that shear forces do not tear the liner 1100 free from thesingle use centrifuge structure 1000. In some instances, an existingmultiple use structure 3000 may be retrofitted for single use processingby selecting a conforming single use structure 1000. In other instances,the multiple use structure 3000 may be specially designed for use withsingle use structure inserts 1000.

FIG. 2 shows a portion of an exemplary structure for upper flanges 1300,plastic liner 1100, and the cover 3200 of a multiple use centrifugestructure 3000 to illustrate sealing the flexible liner 1100 to theupper flanges 1300. The flexible liner 1100 may be a thermoplasticelastomer such as a polyurethane (TPU) or other stretchable, tough,non-tearing, bio-compatible polymer, while the upper and lower flanges1300, 1200 may be fabricated from a rigid polymer such aspolyetherimide, polycarbonate, or polysulfone. The flexible liner 1100is a thin sleeve, or envelope, which extends between and is sealed tothe upper and lower flanges 1300, 1200, and forms the outer wall ofseparation chamber 1550. The composition of the liner 1100 and of theupper and lower flanges 1300, 1200, and core 1510 described herein areexemplary only. Those skilled in the art may substitute suitablematerials with properties similar to those suggested which are, or maybecome, known.

A thermal bonding attachment process may be used to bond the dissimilarmaterials in the area shown in FIG. 2. The thermal bond 1110 is formedby preheating the flange material, placing the elastomeric polymer atopthe heated flange, and applying heat and pressure to the elastomericfilm liner 1100 at a temperature above the film's softening point. Theplastic liner 1100 is bonded to lower flange 1200 in the same manner.Although a thermal bond 1110 is described herein, it is merelyexemplary. Other means of creating a similarly strong relativelypermanent bond between the flexible film and the flange material may besubstituted, such as by temperature, chemical, adhesive, or otherbonding means.

The exemplary single-use components are pre-sterilized. During theremoval of these components from their protective packaging andinstallation into a centrifuge, the thermal bonds 1110 maintainsterility within the single-use chamber. The stretchable flexible liner1100 conforms to the walls of reusable bowl 3100 when in use. Reusablebowl 3100 provides sufficient support, and the flexible liner 1100 issufficiently elastic, to permit the single use structure 1000 towithstand the increased rotational forces which are generated when thelarger radius centrifuge 1000 is filled with a liquid cell culture orother cell suspension and is rotated with a sufficient angular velocityto reach a settling velocity that permits processing at a rate of about2-40 liters a minute.

In addition to the thermal bond 1110, sealing ridges or “nubbins” 3210may be present on bowl cover 3200 to compress the thermoplasticelastomeric film against the rigid upper flanges 1300, forming anadditional seal. The same compression seals may also be used at thebottom of the bowl 3100 to seal the thermoplastic elastomeric filmagainst the rigid lower flanges 1200. These compression seals supportthe thermal bonded areas 1110, by isolating them from shearing forcescreated by the hydrostatic pressure that develops during centrifugationwhen the chamber is filled with liquid. The combination of the thermalbond 1110 and the compression nubbin 3210 seals has been tested at3000×g, which corresponds to a hydrostatic pressure of 97 psi at thebowl wall. The lining should be sufficiently thick and compressible topermit the nubbins 3210 to compress and grip the flexible liner 1100 yetminimize the risk of tearing near the thermal bond 1110 or compressionnubbins 3210. In one embodiment, a flexible TPU liner 0.010 inch thicksealed without tearing or leaking.

An embodiment corresponding to the illustrations of FIGS. 1-2 has beentested within a bowl that was 5.5 inches in diameter. At 2000×g it had ahydraulic capacity>7 liters/min and successfully separated mammaliancells to 99% efficiency at a rate of 3 liter/min.

In most instances, the upper and lower flanges 1300, 1200 have a shapesimilar to that illustrated FIG. 1, but in some instances the uppersurface of the single use centrifuge structure may have a differentshape, as is illustrated in FIGS. 10 and 11. In the embodimentsillustrated in FIGS. 10 and 11, rather than having a generally conicalbowl cover 3200, to conform to generally conical upper flanges 1300,both the upper flanges and the bowl cover are relatively disk-shaped.Those skilled in the art will be able to adapt the sealing techniquesdescribed herein for use with differently shaped sealing surfaces.

FIGS. 4-5 illustrate an embodiment with features to improve theefficiency of the centripetal pump 1400. As shown in detail in FIG. 5,this embodiment of an internal structure for single use componentssimilar to that illustrated in FIGS. 1 and 2 includes a plurality ofradial fins 1630 on the inner face 1620 of a cap portion 1610 of thepump chamber 1420. FIG. 5 shows the inner face 1620 of the cap portion1610 of centrifuge cover 1600. The radial fins 1630, may be thin,generally rectangular, radial plates, extending perpendicularly from theinner surface 1620 of the cap portion 1610. In the exemplary embodiment,six (6) fins 1630 are illustrated, but other embodiments may includefewer or more fins 1630. In this embodiment, fins 1630 form part of theinner face of cap 1620, but in other embodiments may comprise the uppersurface 1620 of pump chamber 1420, which may take a form other than cap1610. When the centrifuge system 1000 is in use, fins 1630 are locatedabove the paring disks 1410 of the centripetal pump 1400 in the chamber1420. These fins 1630 transmit the angular rotation of the centrifuge1000 to the centrate within in the pump chamber 1420.

This increases the efficiency of the centripetal pump 1400, stabilizingthe gas to liquid interface in the pump chamber 1420 above the paringdisks 1410, and increasing the size of the gas barrier. The gas barrieris a generally cylindrical column of gas extending from the exterior ofthe feed/discharge mechanism 2000 outward into the pump chamber 1420 tothe inner surface of the rotating centrate. This increase in the size ofthe barrier occurs because the resulting increase in angular velocity ofthe centrate forces the centrate toward the wall of the centrifuge. Whenrotating centrate within the pump chamber 1420 comes into contact withthe stationary paring disks 1410 the resulting friction may decrease theefficiency of the pump 1400. The addition of a plurality of radial fins1630, which rotate with the same angular velocity as the centrate,overcomes any reduction in velocity that might otherwise result from theencounter between the rotating centrate and the stationary paring disks1410.

FIG. 6 shows an exemplary embodiment of an improved core structure 1500for use in high turbidity feeds. Core structure 1500 includes a core1510, upper flange 1300, and lower flange 1200. Core 1510 has acylindrical central cavity 1520 adapted to permit feed tube 2100 to beinserted into the central cavity 1520. The distance from the centralaxis 1525 to the exterior of core 1515 (the core width, represented bydashed line 6000 in FIG. 6) is larger than the corresponding distance inthe embodiment illustrated in FIG. 3. The larger diameter core 1510decreases the depth (represented by dashed line 6010) of the separationchamber 1550, making centrifuge 1000 operate as a shallow poolcentrifuge. The depth 6010 of a separation chamber 1550 is generally thedistance between the exterior of the core 1510 and the flexible liner1100, labeled in FIGS. 1 and 12. A shallow pool centrifuge is one whichhas a depth 6010 which is small, relative to the diameter of thecentrifuge. As can be seen in the exemplary embodiment illustrated inFIG. 12, in order to facilitate removal of the cell concentrate, theshallow pool depth 6010 may vary from shallower at the bottom ofseparation chamber 1550 to somewhat deeper the top of the separationchamber 1550. In the embodiments illustrated herein, the ratio of theaverage separation pool depth 6010 to the core width is 1:1 or lower. Anexample of a shallow pool centrifuge is offered as an optional model ofthe ViaFuge centrifuge system, manufactured by Pneumatic ScaleCorporation. The advantage of a shallow pool centrifuge is that itenables separation at higher feed flow rates. This is accomplished byvirtue of a higher average g-force for a given inner bowl diameter,which creates a higher sedimentation velocity at a given angularvelocity. The resulting enhanced separation performance is beneficialwhen separating highly turbid feeds containing a high concentration ofcell debris.

The embodiment of the core structure 1500 which is illustrated in FIG. 6also includes accelerator vanes 1560 as part of the lower flange 1200.Accelerator vanes 1560 (as shown in FIG. 12), rather than holes 1530through a solid core 1510 (as shown in FIG. 10-11), comprise analternate embodiment of a fluid connection between the central cavity1520 of the core 1510 and the separation chamber 1550.

In the exemplary embodiment of a core structure 1500 shown in FIG. 6,accelerator vanes 1560 comprise a plurality of radially, generallyrectangular, spaced thin plates 1580 extending upward from the upperconical surface of the lower flange 1200. Plates 1580 extend upwardorthogonal to the base of the core 1510. Plates 1580 generally extendradially outward from near the axis 1525 of the core 1510. In theexemplary embodiment, there are 12 plates 1580, as shown most clearly inFIG. 7. In other embodiments there may be fewer or more than 12 plates1580. In addition, in other embodiments the plates 1580 may be curved inthe direction of rotation of the centrifuge 1000, as shown in anexemplary embodiment in FIG. 9. The interior surface of lower flange1200 may be modified to form an elliptical accelerator bowl 1590, withthe curved plates extending upward therefrom. These embodiments areintended to be exemplary, and those skilled in the art may combine themin different ways or may modify these embodiments to further benefitfrom the turbidity reduction these plates and the shape of the lowerflange 1200 and/or an embedded accelerator bowl create.

Further features of an embodiment of a single use centrifuge 1000 whichis designed to operate continuously or semi-continuously are illustratedin FIGS. 10-12. The exemplary embodiment illustrated in FIG. 10 includesa second centripetal pump 4400 for removal of cell concentrate.Centripetal pump 4400 for the removal of cell concentrate is locatedabove the centripetal pump 1400 for removal of centrate. Centripetalpump 4400 includes a pump chamber 4420 and paring disks 4410. Aplurality of holes or continuous slits 4540 extend from the upper outercircumference of the separation chamber 1550 into pump chamber 4420,providing fluid connection from outer portion of the separation chamber1550 to the second pump chamber 4420. As with pump chamber 1400, pumpchamber 4400 may have a different shape than that illustrated in FIGS.10-12, but will generally be an axially symmetric chamber near the upperend of the core structure 1500 which is in fluid connection with theseparation chamber 1550. As with pump chamber 1400, pump chamber may bepartially or entirely recessed within core structure 1500. If a centratepump chamber 1400 is present near the upper end of the core structure1500, the cell concentrate pump chamber 4400 will generally be locatedabove it. A pump chamber 4400, for the removal of cell concentrate, willbe in fluid connection with separation chamber 1550 via holes or slits4540 which extend from adjacent the outer upper wall of separationchamber 1550, in order to collect the heavier cell concentrate which isurged there by centrifugal forces.

In the embodiment illustrated in FIG. 10, the paring disks 4410 used inthe concentrate discharge pump 4400 are approximately the same radius asthose used in the centrate discharge pump 1400, and are rotationallyfixed. In other embodiments, such as the one shown in FIG. 11, theparing disks 4410 in the concentrate discharge pump 4400 may have alarger radius than those in the centrate discharge pump 1400, with acorrespondingly larger pump chamber 4420. Paring disks of variousintermediate diameters may be used as well. The optimum diameter willdepend on the properties of the cell concentrate that is to bedischarged. Larger diameter paring disks have a higher pumping capacity,but create greater shear.

In the embodiments illustrated in FIGS. 1, 4, and 10, the paring disks4410 in the concentrate discharge pump 4400 are rotationally fixed. Inother embodiments, such as the one shown in FIG. 11, paring disks in4410 may be adapted to rotate with an angular velocity between zero andthe angular velocity of the centrifuge 1000. The desired angularvelocity can be controlled by a number of mechanisms that are known tothose skilled in the art. An example of a means of control is anexternal slip clutch that allows the paring disks 4410 to rotate at anangular velocity that is a fraction of that of the centrifuge 1000.Other means of controlling the angular velocity of the paring disks willbe apparent to those skilled in the art.

In the embodiments illustrated in FIGS. 1, 4, 10-12, the gaps 1415, 4415between paring disks 1410 and 4410 are fixed. In other embodiments, suchas the embodiment in FIG. 13, the gaps 1415, 4415 between paring disks1410 and 4410 may be adjustable, in order to control the flow rate atwhich centrate or concentrate are removed from the centrifuge 1000. Oneof each pair of paring disks 1410 and 4410 is attached to a verticallymoveable throttle tube 6100. Throttle tube 6100 may be moved up or downin order to narrow or widen the gap 1415, 4415 between each pair of theparing disks 1410, 4410. In addition, an external peristaltic pump 2510(not shown) may be added to the concentrate removal line 2500 (notshown) to aid in removal of concentrate. This pump 2510 may becontrolled by a sensor 4430 in the pump chamber 4420. The sensor 4430(not shown) may also be used to control a diluent pump 5150 in order tosynchronize concentrate removal with the addition of diluents.

Also illustrated in FIG. 13 is an embodiment in which the centrate pump1400 is located at the base of the centrifuge 1000. In the embodimentillustrated in FIG. 13, a centrate well 1555 is created between the pumpchamber 1420 and the flexible liner 1100. Holes 1530 extend from thecore 1510, below the pump chamber 1420, into the centrate well 1555. Inaddition, in the exemplary embodiment illustrated, holes 1540 extendfrom the separation chamber 1550, adjacent the exterior surface 1515 ofthe core 1510, into the pump chamber 1420 to permit the centrate to beremoved using centrate pump 1400. Holes 4540 may also extend between theseparation chamber 1550, adjacent its outer upper surface, into pumpchamber 4420 to permit cell concentrate to flow into pump chamber 4420to be removed using centripetal pump 4400.

As noted above, in the exemplary embodiment illustrated, the gaps 1415,4415 between the paring disks 4410 and 1410 may be adjustable by use ofa throttle tube 6100 connected to one of each pair of paring disks 4410,1410. Throttle tube 6100, and the attached one of each paring disk pair4410, 1410, may be moved up or down to narrow or widen gaps 1415, 4415.In the exemplary embodiment illustrated, the throttle tube 6100 isattached to the lower and upper paring disk of paring disk pairs 4410,1410, respectively. In other embodiments the attachment may be reversed,may be used to throttle a single centripetal pump, or may be used tothrottle both in parallel (rather than opposition as illustrated in FIG.13).

As can be seen in the embodiments illustrated in FIGS. 10-12, the wallof the solid multiple use bowl 3100 is thicker at the base than it is inthe upper portion, in order to create an internal truncated cone shapeto support single use centrifuge structure 1000 which has a smallerradius at the lower end than at the upper end. This larger radius at theupper end of the separation chamber 1550 moves the denser cellconcentrate toward the upper outer portion of the separation chamber1550 and into centripetal pump chamber 4420. In the embodimentillustrated, the truncated cone shape is created by a multiple use bowl3100 with a wall which is thicker at the base than it is in the upperportion. Those skilled in the art will recognize that a multiple usebowl 3100 having an internal truncated cone shape may also include wallsof uniform thickness, and that there may be other variations whichcreate the desired internal shape for the multiple use bowl 3100.

In the embodiments illustrated in FIGS. 10-12, feed mechanism 2000 alsoincludes an additional pathway for the removal of cells, or cellconcentrate. In the embodiment illustrated in FIG. 1, the cylindricalpathway 2200 around the feed tube 2100 is used to remove centrate. Theembodiments illustrated in FIGS. 10-12 also include, a concentriccylindrical pathway for the removal of cells or cell concentrate,referred to as a cell discharge tube 2500. Cell discharge tube 2500surrounds the centrate removal pathway 2200. If the centrifuge isdesigned to be used with a concentrate that is expected to be veryviscous, an additional concentric cylindrical fluid pathway 5000 may beadded around the feed tube 2100 to permit the diluents to be introducedinto the cell concentrate pump chamber 4420 in order to decrease theviscosity of the concentrate. The diluent pathway 5000, in the exemplaryembodiment illustrated in FIG. 12, comprises a concentric tubesurrounding the cell discharge pathway, and opens at the lower end intoa thin disk-shaped fluid pathway 5100 above paring disks 4410,discharging near the outer edge of the paring disks 4410 to providefluid communication with the pump chamber 4420. Injecting the diluent bythis means, and in this location, limits the diluent to mixing with, andbeing discharged with, the concentrate rather than being introduced intothe centrate, which may be undesirable in some applications. Inalternative embodiments, the diluents may be introduced directly ontothe upper surface of the paring disks and allowed to spread radiallyoutward, or onto a separate disk located above the paring disks.

The choice of diluent will depend on the objectives of the separationprocess and the nature of the cell concentrate that is to be diluted. Insome cases a simple isotonic buffer or deionized water can serve as thediluent. In other cases, diluents that are specific to the properties ofa cell concentrate may be advantageous. For example, in production scalebatch cell culture operated at low cell viability, flocculants arecommonly added to the culture as it is being fed to a centrifuge tocause cells and cell debris to flocculate or agglomerate into largerparticles, which facilitates their separation by increasing their rateof sedimentation. Since both cells and cell debris carry negativesurface charges, the compounds used as flocculants are typicallycationic polymers, which carry multiple positive charges, such aspolyethyleneimine. By virtue of their multiple positive charges, suchflocculants can link negatively charged cells and cells debris intolarge agglomerates. An undesirable consequence of the use of suchflocculants is that they further increase the viscosity of cellconcentrates. Therefore, a particularly useful diluent in thisapplication is a deflocculant that will disrupt the bonds that increasethe viscosity of the cell concentrate. Examples of deflocculants includehigh salt buffers such as sodium chloride solutions ranging inconcentration from 0.1 M to 1.0 M. Other deflocculants that may beuseful in reducing the viscosity of cell concentrate are anionicpolymers such as polymers of acrylic acid.

In the case of a cell concentrate wherein cell viability is to bemaintained, a diluent can be chosen that is a shear protectant, such asdextran or Pluronic F-68. The use of a shear protectant, in combinationwith an isotonic buffer, will enhance the survival and viability ofcells as they are being discharged from the centrifuge.

The exemplary centrifuge illustrated in FIG. 4 operates as describedbelow. During a feed cycle, a feed suspension flows into the rotatingbowl assembly through feed tube 2100. As the feed suspension enters thecentral cavity 1520 of core 1510 near lower flange 1200, it is urgedoutward along the upper surface of lower flange 1200 by centrifugalforces, passing into the separation chamber 1550 through holes 1530 incore 1510.

Centrate collects in the separation chamber 1550, a hollow, roughlycylindrical space below the upper flange 1300 surrounding core 1510. Thecentrate flows upward from its entrance into the separation chamberthrough holes 1530 until it encounters holes 1540 between the separationchamber 1550 and the pump chamber 1420 in the upper portion of theseparation chamber 1550, adjacent the core 1410. Particles of densityhigher than that of the liquid are moved toward the outer wall of theseparation chamber 1550 by sedimentation (particle concentrate), awayfrom holes 1530. When the rotation of the centrifuge 1000 is stopped,the particle concentrate moves downward under the influence of gravityto the nozzle 2110 of the feed tube 2100 for removal via the combinedfeed/discharge mechanism 2000.

During rotation, the centrate enters the centrate pump chamber 1420through holes 1540. Within the pump chamber 1420, the rotating centrateencounters stationary paring disks 1410, which convert the kineticenergy of the rotating liquid into pressure which urges the centratebeing discharged upward through the centrate discharge path 2200 withinthe feed/discharge mechanism 2000 and out through the centrate dischargetube 2400.

The efficiency of the centripetal pump 1400 is increased by addingradial fins 1630 on the inner surface 1620 of the cap portion 1610 ofthe rotating pump 1400. These fins 1630 impart the angular momentum ofthe rotating assembly to the centrate in the pump chamber 1420, whichmight otherwise slow because of friction when the rotating centrateencounters the stationary paring disks 1410. The centripetal pump 1400provides an improved means of centrate discharge, over mechanical seals,because of the gas liquid interface within the pump chamber 1420. Thegas within the pump chamber 1420 is isolated from contamination by theexternal environment by the rotating seal 1700. Because the centratebeing discharged between the paring disks 1410 does not come intocontact with air, either during the feed or discharge process, it avoidsthe excessive foaming that often occurs when the discharge processintroduces air into the cell culture.

In the centrifuge 1000 embodiment illustrated in FIGS. 4-5, cellconcentrate is discharged by periodically stopping bowl rotation and thefeed flow and then pumping out the cell concentrate that has beencollected along the outer walls of the separation chamber 1550. Thisprocess is known as intermittent processing. When the volumetriccapacity of the separation chamber 1550 is reached, centrifuge rotationis stopped. The cell concentrate moves downward toward nozzle 2110 offeed tube 2100, where the concentrate is withdrawn by pumping it outthrough the feed tube 2100. Appropriate valving (not shown) external tothe centrifuge 1000 is used to direct the concentrate into a collectionvessel (not shown). If the entire bioreactor batch has not yet beencompletely processed, then bowl rotation and feed flow are resumed, andis followed by additional feed and discharge cycles until the full batchhas been processed.

As noted above, when the cell culture is concentrated or containssignificant cell debris, the process described above slows down becauseresidence time must be increased to capture small debris particles,which necessitates a slower feed flow rate and the separation chamber1550 fills rapidly and rotation must be halted frequently and repeatedlyfor each culture batch. In addition, the cell concentrate tends to bemore viscous so gravity does not work as efficiently to drain the cellconcentrate to the bottom of the centrifuge 1000 so it takes longer and,in some instances, may require a wash to remove the remaining cells.

The single use centrifuge, as modified in the exemplary embodimentsillustrated in FIGS. 6-13, creates a higher average settling velocitywithout an increase in angular velocity, permits the centrifuge 1000 torun continuously or semi-continuously, and allows a diluent to be addedto the cell concentrate during the cell removal process so that theremoval of cells is more easily and more completely accomplished.

Embodiments of a single use centrifuge structure 1000 shown in FIGS.6-12 operate as discussed herein. Feed suspension enters the single usecentrifuge structure 1000 via feed tube 2100. As the feed suspensionencounters accelerator vanes 1560, the vanes 1560 impart an angularvelocity to the feed suspension which approaches the angular velocity ofthe single use centrifuge 1000. The use of vanes 1560, rather than holes1530, provides for a greater volume of feed suspension to enter theseparation chamber 1550 at a slower radial velocity, avoiding thejetting which occurs when the feed suspension is forced through holes1530 having smaller cross sectional openings than the openings betweenthe vanes 1560. This reduction in velocity of the feed stream as itenters the separation zone, or pool, minimizes disruption of the liquidcontents of the pool, which allows for more efficient sedimentation.

As the centrifuge 1000 rotates, the particles which are denser than thecentrate are urged toward the outside of the separation chamber 1550,leaving the particle free centrate near the core 1510. The centrifugebowl 3100 has the shape of an inverted truncated cone, with a widerradius at the upper end than the lower end. The centrifugal force causesthe particles to collect in the upper and outer portion of the chamber.The centrifuge 1000 may operate with semi-continuous discharge ofconcentrate. The centrate discharge works, generally, as described withrespect to FIG. 4. The cell concentrate discharge works similarly, withthe cell concentrate collecting near the upper outer portion of theseparation chamber 1550 and entering the concentrate discharge pumpchamber 4400 via holes 4540 adjacent the upper outer wall of theseparation chamber 1550.

The rate of feed of suspension, as well as the angular velocity ofrotation, may be monitored using a vibration sensor system such as theone described in published PCT Application No. WO 2011/123371,incorporated by reference herein in its entirety. Such a sensor systempermits the centrifuge to be filled at a lower rate until the vibrationsindicate the centrifuge is nearly full, then to adjust the feed rate andangular velocity appropriately in response to this information.Typically, the feed rate will be decreased or stopped once thecentrifuge is nearly full and the angular velocity will be increased inorder to increase the settling velocity and once the settling anddischarge is essentially complete, the cycle will be repeated. If thesystem is optimized using the additional features described herein todiminish the need to interrupt the process, it may be possible tooperate the system continuously, or nearly continuously, at the angularvelocity needed for settling.

With semi-continuous concentrate discharge, the suspension continues tobe fed into the centrifuge 1000, using concentrate pump 4400 operatingintermittently to remove concentrate. The operation of concentrate pump4400 may be controlled by an optical sensor in the concentrate dischargeline that indicates the presence or absence of concentrate beingdischarged. In lieu of a concentrate pump 4400, the discharge cycle maybe managed electronically using a controller and sensors which determinewhen to open and shut a valve for the most efficient processing of thefluid suspension.

The average rate of discharge may further be controlled by using acentrifuge 1000 with an adjustable gap between the paring disks 4410,1410. It should be noted that it may only be desired or necessary forone set of paring disks 4410, 1410 to be adjustable. The gap betweenparing disks 4410, 1410 (which forms a part of the fluid pathway out ofthe centrifuge 1000) may be opened to permit flow, or closed to shut theflow off, acting as an internal valve. Depending on the desired product,or the characteristics of the product, it may also be useful to widen ornarrow the gap 4415, 1415 between paring disks 4410, 1410. Changing thegap affects both pumping and shear rates associated with the pairingdisks.

The rate of removal of concentrate and centrate from the centrifuge1000, and the viability of the concentrate removed, may be furthercontrolled using a number of features of exemplary embodiments shown inFIGS. 4-13. Accelerator fins 4630, similar to those in the centrate pumpchamber 1420, may be added to concentrate pump chamber 4420. Theaddition of accelerator fins 4630 increases the rate at which theconcentrate may be removed, by overcoming some of the slow down due tofriction between the moving concentrate and the paring disks 4410. Inaddition to accelerator fins 4630 in the upper surface of the pumpchamber 4420, such fins 4630 may also be added to a lower surface in thepump chamber 4420 to increase their effectiveness. A further feature maybe the substitution of slits for holes 1540, 4540, which minimizes theshear on material entering the pump chambers 1420, 4420.

If viability of the concentrate is a concern, rotatable paring disks4410 may be included in pump chamber 4420, which reduce the shearimparted to the concentrate as it contacts the surfaces of the paringdisks 4410. The rotation rate of paring disks 4410 may be adjusted to arate somewhat between stationary and the rate of rotation of theseparation chamber 1550 to balance concentrate viability against therate of discharge. The desired angular velocity can be controlled by anumber of mechanisms that are known to those skilled in the art. Anexample of a means of control is an external slip clutch that allows theparing disks to rotate at an angular velocity that is a fraction of thatof the centrifuge. The use of slip clutches is well known to thoseskilled in the art. In addition, there may be means other than slipclutches to adjust the angular velocity that will be apparent to thoseskilled in the art.

A peristaltic pump 2510 may be also used to make removal of theconcentrate more efficient and reliable, particularly with veryconcentrated feed suspensions. Using a peristaltic pump 2510 permits theuser to more precisely control the rate of flow of the concentrate fromthe centrifuge 1000 than is possible relying on centripetal pumps 4400,alone, because the rate of centripetal pumps are not as easilyadjustable as the rate of a peristaltic pump 2510.

In addition, a diluent, such as sterile water or a buffer, may be pumpedinto the concentrate pump chamber 4420 through the diluent pathway 5000using a diluent pump 5150 in order to cut the viscosity of theconcentration. A more complete discussion of useful diluents can befound above. The rate at which either or both of the peristaltic pump2510 or the diluent pump 5150 operates may be controlled by an automatedcontroller (not shown) responsive to a concentration sensor 4430 locatedin the concentrate discharge connection 2500. The controller may beprogrammed to start, stop, or modify the pump rate for both diluentaddition and concentrate removal responsive to the particleconcentration in the concentrate, either independently, responsive to aconcentration sensor 4430, in conjunction with a standard feed/dischargecycle, or as a combination.

FIG. 16 shows an alternative embodiment of a core used in connectionwith a centrifuge that provides continuous separation processing toproduce continuous concentrate and centrate feeds. The core 10 issimilar to those previously discussed that is configured to bepositioned in the rotatable bowl of a centrifuge. The centrifuge bowland the core rotate about an axis 12 during processing. The apparatusincludes a stationary assembly 14 and a rotatable assembly 16.

As with the previously described embodiments, the stationary assembly 14includes a feed tube 18. The feed tube 18 is coaxial with the axis 12and terminates in an opening 20 adjacent the bottom of the separationchamber or cavity 22 of the core. The stationary assembly furtherincludes a centrate centripetal pump 24. The exemplary embodiment of thecentrate centripetal pump 24, which is described in greater detailhereafter, includes inlet opening 26 and an annular outlet opening 28.The annular outlet opening is in fluid connection with a centrate tube30. The centrate tube extends in coaxial surrounding relation with thefeed tube 18.

In this exemplary embodiment, the centrate centripetal pump 24 ispositioned in a centrate pump chamber 32. The centrate pump chamber isdefined by walls which are part of the rotatable assembly, and whichduring operation provide for the inlet openings 26 of the centratecentripetal pump to be exposed to a pool of liquid centrate.

The exemplary embodiment further includes a concentrate centripetal pump34. The concentrate centripetal pump 34 of the exemplary embodiment isalso of a construction like that later discussed in detail. In theexemplary arrangement the concentrate centripetal pump 34 includes inletopenings 36 positioned in a wall that bounds the annular periphery ofthe centripetal pump. It should be noted that the concentratecentripetal pump 34 has a greater peripheral diameter than theperipheral diameter of the centrate pump. The concentrate pump furtherincludes an annular outlet opening 38. The annular outlet opening 38 isin fluid connection with a concentrate outlet tube 40. The concentrateoutlet tube extends in coaxial surrounding relation with the centratetube 30.

In the exemplary embodiment the inlet openings 36 of the concentratecentripetal pump are positioned in a concentrate pump chamber 42. Theconcentrate pump chamber is defined by walls of the rotatable assembly16. During operation, the inlet openings 36 of the concentratecentripetal pump are exposed to concentrate in the concentrate pumpchamber 42. The concentrate pump chamber 42 is bounded vertically by atop portion 44. At least one fluid seal 46 extends between the outercircumference of the outlet tube 40 and the top portion 44. Theexemplary seal 46 is configured to reduce the risk of fluid escapingfrom the interior of the separation chamber and to prevent introductionof contaminants from the exterior area of the core therein.

During operation of the centrifuge, the bowl and the core including thecavity or separation chamber is rotated about the axis 12 in arotational direction. Rotation in the rotational direction is operativeto separate cell suspension that is introduced through the feed tube 18,into centrate which is discharged through the centrate tube 30 andconcentrate which is discharged through the concentrate outlet tube 40.

Cell suspension enters the separation chamber 22 through the tubeopening 20 at the bottom of the separation chamber. The cell suspensionis moved outwardly via centrifugal force and a plurality of acceleratorvanes 48. As the suspension is moved outwardly by the accelerator vanes,the cell suspension material is acted upon by the centrifugal force suchthat the cell material is caused to be moved outwardly toward theannular tapered wall 50 that bounds the outer side of the separationchamber. The concentrated cellular material is urged to move outwardlyand upwardly as shown against the tapered wall 50 and through aplurality of concentrate slots 52. The concentrate material movesupwardly beyond the concentrate slots and into the concentrate pumpchamber 42 from which the concentrate is discharged by the concentratecentripetal pump 34.

In the exemplary arrangement, during operation the cell free centrate ispositioned in proximity to a vertical annular wall 54 which bounds theinside of the separation chamber 22. The centrate material movesupwardly through centrate holes in the annular base structure thatbounds the centrate pump chamber 32. The centrate moves upwardly throughthe centrate holes 56 and forms a pool of liquid centrate in thecentrate chamber. From the centrate chamber, the centrate is movedthrough operation of the centrate centripetal pump 24 and delivered fromthe core through the centrate tube 30.

In the exemplary embodiment of FIG. 16, the concentrate and centratepumps have a configuration generally like that shown in FIG. 17. In FIG.17, the centrate centripetal pump 24 is represented in an isometricexploded view. As shown in FIG. 17, the exemplary centripetal pump has adisk-shaped body that is comprised of a first plate 58 and a secondplate 60. During operation, the first plate and the second plate areheld in releasible engaged relation via fasteners which are representedby screws 62. Of course it should be understood that in otherembodiments, other configurations and fastening methods may be used.

In the exemplary arrangement, the second plate 60 includes walls thatbound three sides of curved volute passages 64. It should be understoodthat while in the exemplary arrangement, the centripetal pump includes apair of generally opposed volute passages 64. In other arrangements,other numbers and configurations of volute passages may be used.

In the exemplary arrangement, the first and second plates make up thedisk-shaped body of the centripetal pump which has a annular verticallyextending wall 67 which defines an annular periphery 66. Inlet openings68 to the volute passages 64 extend in the annular periphery. An annularcollection chamber 70 extends in the body radially outwardly from theaxis 12 and is fluidly connected to the volute passages. The annularcollection chamber 70 receives the material that enters the inletopenings 68. The annular collection chamber 70 is in fluid connectionwith an annular outlet opening that is coaxial with the axis 12. In theexemplary arrangement for the centrate centripetal pump, the annularoutlet opening is an annular space which extends between the outer wallof feed tube 18 and the inner wall of second plate 60 which outlet isfluidly connected to the centrate outlet tube 30.

In the exemplary arrangement each of the volute passages 64 isconfigured such that the volute passages are curved toward therotational direction of the bowl and separation chamber, the rotationaldirection is represented by Arrow R in FIG. 17. In the exemplaryarrangement, the vertically extending walls 74 which bound the volutepassages and which face the rotational direction, are each curved towardthe rotational direction. The curved configuration of the walls 74 whichbound the volute passages horizontally, provide for the enhanced pumpingproperties of the exemplary arrangement. Further, the opposed boundingwall 76 of each volute passage in the exemplary arrangement has asimilar curved configuration. The curved configuration of the verticallyextending walls that bound the volute passages horizontally provide fora constant cross-sectional area of each volute passage from therespective inlet to the collection chamber. This consistentcross-sectional area is further achieved through the use of a generallyflat wall 78 which extends between walls 74 and 76 and which bounds thevolute passage vertically on one side. Further in the exemplaryembodiment the first plate 58 includes a generally planar circular face80 on its side which faces inwardly when the plates are assembled toform the disk-shaped body of the centripetal pump. In this exemplaryarrangement, the face 80 serves to vertically bound the sides of bothvolute passages 64 of the centripetal pump.

Of course it should be understood that this exemplary arrangement whichincludes a pair of plates, one of which includes a recess with wallswhich bound three of the four sides of the curved volute passages andthe other of which includes a surface that bounds the remaining side ofthe volute passages is exemplary. It should be understood that in otherarrangements, other configurations and structures may be used.

In the exemplary centripetal pump structure shown in FIG. 16, thecentripetal pump structures are utilized and have the capability formoving more liquid than comparably sized paring disk-type centripetalpumps. Further, the exemplary configuration produces less heating of theliquid than comparable paring disks.

Further in the exemplary arrangement as previously discussed, theannular periphery of the centrate centripetal pump 24 has a smallerouter diameter than the periphery of the concentrate centripetal pump34. This configuration is used in the exemplary arrangement to avoid thecentrate centripetal pump removing too much liquid from the pool ofliquid centrate which forms in the centrate pump chamber 32. Assuringthat there is sufficient liquid centrate within the centrate pumpchamber, helps to assure that waves do not form in the centrate adjacentto the inlets of the centrate centripetal pump. The formation of waveswhich could result from less than sufficient liquid centrate, may causevibration and other undesirable properties of the centrifuge and core.

The larger annular periphery of the concentrate pump of the exemplaryarrangement causes material to preferentially flow out of the core viathe concentrate centripetal pump. In exemplary arrangements, the flow ofconcentrate downstream of the concentrate output tube 40 can becontrolled to control the ratio of centrate flow to concentrate flowfrom the core.

Further in exemplary embodiments, utilizing centripetal pumps having theconfigurations described, the properties and flow characteristics of thecentrifuge may be tailored to the particular materials and requirementsof the separation processing being performed. Specifically the diametersof the annular periphery of the centripetal pumps may be sized so as toachieve optimum properties for the particular processing activity. Forexample, the larger the diameter of the periphery of the centripetalpump, the greater flow and pressure at the outlet that can be achieved.Further the larger diameter tends to produce greater mixing than arelatively smaller diameter. However, the larger diameter also resultsin greater heating than a smaller peripheral diameter of a centripetalpump. Thus to achieve less heating, a smaller diameter periphery may beused. Further it should be understood that different sizes, areas andnumbers of inlet openings and different volute passage configurationsmay be utilized to vary flow and pressure properties as desired forpurposes of the particular separation process.

FIG. 19 shows schematically an exemplary system which is utilized tohelp assure positive pressure within a separation chamber which isalternatively referred to herein as a cavity, during cell suspensionprocessing. As discussed in connection with previous exemplaryembodiments, it is generally desirable to assure positive pressure aboveatmospheric pressure at all times within the separation chamber. Doingso reduces the risk that contaminants are introduced into the separationchamber by infiltrating past the one or more fluid seals which extendbetween the stationary assembly and the rotatable assembly of the core.Further as previously discussed, it is also generally desirable tomaintain air at positive pressure within the separation chamber incontact with the interior face of the fluid seal. The presence of an airpocket adjacent the seal avoids the seal coming into contact with thematerial being processed and further helps to reduce the risk ofcontaminant introduction into the processed material as well as theescape of any material from the separation chamber.

The exemplary system described in connection with FIG. 19 serves tomaintain a consistent positive pressure in the separation chamber andreduces the risk of the introduction of contaminants and the escape ofprocessed material.

As schematically shown in FIG. 19, the centrifuge includes a rotatablebowl 82. The centrifuge bowl is rotatable about an axis 84 by a motor 86or other suitable rotating device.

The exemplary centrifuge structure shown includes a rotatable single usecore 88 which bounds a cavity 90 which is alternatively referred toherein as a separation chamber.

Like other previously described embodiments, the exemplary core includesa stationary assembly which includes a suspension inlet feed tube 92which has an inlet opening 94 positioned adjacent to the bottom area ofthe cavity. The stationary assembly further includes at least onecentripetal pump 96. The centripetal pump of the exemplary embodimentincludes a disk-shaped body with at least one pump inlet 98 adjacent theperiphery thereof and a pump outlet 100 adjacent the center of thecentripetal pump. The pump outlet is in fluid connection with a centrateoutlet tube 102. The centrate outlet tube extends in coaxial surroundingrelation of the suspension inlet tube in a manner similar to thatpreviously discussed. The rotatable top portion 104 of the fluidcontaining separation chamber is in operative connection with at leastone seal 106. The at least one seal 106 extends operatively in sealingrelation between the outer annular surface of the centrate outlet tube102 which is stationary, and the rotatable top portion 104 of the core.

In the exemplary arrangement the inlet tube 92 is fluidly connected to apump 108. Pump 108 in an exemplary arrangement is a peristaltic pumpwhich is effective to pump cell suspension without causing damagethereto. Of course it should be understood that this type of pump isexemplary and in other arrangements, other types of pumps may be used.Further in the exemplary arrangement the pump 108 is reversible. Thisenables the pump 108 to act as a feed pump so as to be able to pump cellsuspension from an inlet line 110 and into the inlet tube at acontrolled rate. Further in the exemplary arrangement, the pump 108 mayoperate as a concentrate removal or discharge pump after the cellconcentrate has been separated by centrifugal action. In performing thisfunction, the pump 108 operates to pump cell concentrate out of theseparation chamber by reversing the flow of material in the inlet tube92 from that when cell suspension is fed into the separation chamber.The cell concentrate is then pumped to a concentrate line 112. Asrepresented in FIG. 19, the inlet line 110 and concentrate line 112 canbe selectively opened and closed by valves 114 and 116 respectively. Inthe exemplary embodiments, valves 114 and 116 comprise pinch valveswhich open and close off flow through flexible lines or tubing. Ofcourse it should be understood that this approach is exemplary and inother arrangements, other approaches may be used.

In the exemplary system, the centrate outlet tube 102 is fluidlyconnected to a centrate discharge line 118. The centrate discharge lineis fluidly connected to a centrate discharge pump 120. In the exemplaryarrangement, the centrate discharge pump 120 is a variable flow ratepump which can have the flow rate thereof selectively adjusted. Forexample in some exemplary arrangements, the pump 120 may include aperistaltic pump which includes a motor, the speed of which may becontrolled so as to selectively increase or decrease the flow ratethrough the pump. The outlet of the centrate discharge pump delivers theprocessed centrate to a suitable collection chamber or other processingdevice.

In the exemplary arrangement schematically represented in FIG. 19, apressure damping reservoir 122 is fluidly connected to the centratedischarge line 118 fluidly intermediate of the centrate outlet tube 102and the pump 120. In the exemplary arrangement, the pressure dampingreservoir includes a generally vertically extending vessel with aninterior area configured for holding liquid centrate in fluid tightrelation. The pressure damping reservoir includes a bottom port 124which is fluidly connected to the centrate discharge line 118.

On an opposed side of the reservoir 122 is a top port 126. The top portis exposed to air pressure. In the exemplary arrangement, the top portis exposed to air pressure from a source of elevated air pressureschematically indicated 128. In exemplary embodiments, the source ofelevated pressure may include a compressor, an air storage tank or othersuitable device for providing a source of elevated air pressure aboveatmospheric pressure within the range needed for operation of thesystem. Air from the source of elevated pressure 128 is passed through asterile filter 130 to remove impurities therefrom. A regulator 132 isoperative to maintain a generally constant air pressure level aboveatmospheric at the top port of the pressure damping reservoir. Inexemplary arrangements, the air pressure regulator comprises anelectronic fast acting regulator to help assure that the generallyconstant air pressure at the desired level is maintained. The exemplaryfast acting regulator 132 operates to rapidly increase the pressureacting at the top port 126 when the pressure falls below the desiredlevel, and relieves pressure rapidly through the regulator in the eventthat the pressure acting at the top port is above the set value of theregulator.

In some embodiments the regulator outlet may also be in operative fluidconnection with the interior of the top portion 104 of the separationchamber through an air line 143 shown schematically in phantom. In suchexemplary arrangements the outlet pressure of the regulator that acts onthe top port 126 of the reservoir also acts through the air line 143 onthe air pocket inside of the separation chamber above the centripetalpump inlet and on the interior of the at least one seal 106. In theexemplary arrangement the line 143 applies the positive pressure to thearea within the separation chamber below the at least one seal throughat least one segregated passage that extends through the stationarystructures of the assembly which includes the centrate outlet tube 102and the inlet feed tube 92. The at least one exemplary segregatedpassage of the air line 143 applies the air pressure to the interior ofthe top portion 104 through at least one air opening 145 to theseparation chamber. The exemplary at least one opening 145 is positionedoutside the exterior surface of the outlet tube 102, above the inlet 98to the centripetal pump and below the at least one seal 106. Of courseit should be understood that this described structure for the exemplaryair line that provides positive air pressure to the air pocket in theseparation chamber and on the inner side of the at least one seal isexemplary, and in other embodiments, other structures and approaches maybe used.

In the exemplary arrangement of the pressure damping reservoir 122, anupper liquid level sensor 134 is configured to sense liquid centratewithin the interior of the pressure damping reservoir. The upper liquidlevel sensor is operative to sense liquid at an upper liquid level. Alower liquid level sensor 136 is positioned to sense liquid in thereservoir at a lower liquid level. A high liquid level sensor 138 ispositioned to detect a high liquid level in the reservoir above theupper liquid level. The high liquid level sensor is positioned so as tosense a liquid level at an unacceptably high level so as to indicate anabnormal condition which may require shutting down the system or takingother appropriate safety actions. In the exemplary arrangement, theliquid level sensors 134, 136 and 138 comprise capacitive proximitysensors which are suitable for sensing the level of the liquid centrateadjacent thereto within the pressure damping reservoir. Of course itshould be understood that these types of sensors are exemplary and inother arrangements, other sensors and approaches may be used.

The exemplary embodiment further includes other components as may beappropriate for the operation of the system. This may include othervalves, lines, pressure connections or other suitable components forpurposes of carrying out the processing and handling of the suspension,centrate and concentrate as appropriate for the particular system. Thismay include additional valves such as valve 140 shown schematically forcontrolling the open and closed condition of the centrate discharge line118. The additional lines, valves, connections or other items includedmay vary depending on the nature of the system.

The exemplary system of FIG. 19 further includes at least one controlcircuit 142 which may be alternatively referred to as a controller. Theexemplary at least one control circuit 142 includes one or moreprocessors 144. The processor is in operative connection with one ormore data stores schematically indicated 146. As used herein, aprocessor refers to any electronic device that is configured to beoperative via processor executable instructions to process data that isstored in the one or more data stores or received from external sources,to resolve information, and to provide outputs which can be used tocontrol other devices or carry out other actions. The one or morecontrol circuits may be implemented as hardware circuits, software,firmware or applications that are operative to enable the controlcircuitry to receive, store or process data and to carry out otheractions. For example the control circuits may include one or more of amicroprocessor, CPU, FPGA, ASIC or other integrated circuit or othertype circuit that is capable of performing functions in the manner of anelectronic computing device. Further it should be understood that datastores may correspond to one or more of volatile or nonvolatile memorydevices such as RAM, flash memory, hard drives, solid state devices,CDs, DVDs, optical memory, magnetic memory or other circuit readablemediums or media upon which computer executable instructions and/or datamay be stored.

Circuit executable instructions, may include instructions in any of aplurality of programming languages and formats including, withoutlimitation, routines, subroutines, programs, threads of execution,objects, methodologies and functions which carry out the actions such asthose described herein. Structures for the control circuits may include,correspond to and utilize the principles described in the textbookentitled Microprocessor Architecture, Programming, and Applications withthe 8085 by Ramesh S. Gaonker (Prentiss Hall, 2002), which isincorporated herein by reference in its entirety. Of course it should beunderstood that these control circuit structures are exemplary and inother embodiments, other circuit structures for storing, processing,resolving and outputting information may be used.

In the exemplary arrangement, the at least one control circuit 142 is inoperative connection through suitable interfaces with at least onesensor such as sensors 134, 136 and 138. The at least one controlcircuit is also in operative connection with the variable flow ratedischarge pump 120. Further in some exemplary embodiments, the at leastone control circuit may also be in operative connection with otherdevices such as motor 86, pump 108, regulator 132, air pressure source128, the fluid control valves and other devices.

The exemplary at least one control circuit is operative to receive dataand control such devices in accordance with circuit executableinstructions stored in the data store 146. In the exemplary arrangement,the fluid level 147 in the fluid damping reservoir is a property thatcorresponds to pressure in the centrate discharge tube 102. In oneexemplary implementation which does not utilize air line 143, the factthat the pressure in the centrate discharge tube is indicative of thepressure in the top portion 104 of the core and the nature of thepressure in the separation chamber adjacent to the seal 106 is utilizedto control the operation of the discharge pump and other components. Aspreviously discussed, it is desirable to maintain a positive pressureabove atmospheric pressure and a pocket of air adjacent to the at leastone seal within the separation chamber to avoid the introduction ofcontaminants into the separation chamber which could result fromnegative pressure. However, if the fluid level becomes too high withinthe separation chamber, the pressure and the suspension material beingprocessed may overflow the seal which may result in potentialcontamination and undesirable exposure and loss of processed material.This may result from conditions where the back pressure on the centrateline which is in connection with the outlet from the centripetal pump istoo high.

In the exemplary arrangement the bowl speed produces a correspondingpumping force and a pump output pressure level of the centripetal pump.This pump output pressure level of the centripetal pump varies with therotational speed of the bowl and the core. The exemplary arrangementwithout the use of air line 143 provides for a back pressure to becontrolled on the centrate outlet tube. Back pressure is provided bycontrolling the speed of a motor operating the pump 120 and the liquidlevel 147 in the pressure damping reservoir. The back pressure ismaintained so as to be less than the pump output pressure level (so thatthe centripetal pump may deliver centrate out of the separation chamber)but is maintained at a positive pressure above atmospheric so as toassure that contaminants will not infiltrate into the separation chamberpast the seal, and so that air at elevated pressure is maintained in theinterior of the separation chamber adjacent to the seal so as to isolatethe seal from the components of the suspension being processed.

In the exemplary arrangement the elevated pressure applied to the topport 126 of the pressure damping reservoir is maintained by theregulator 132. Further by the at least one control circuit 142controlling the speed of pump 120 to maintain the liquid level 147between the upper liquid level as sensed by the sensor 134 and the lowerliquid level 136, centrate flow out of the separation chamber iscontrolled so that the pressure of the top area of the separationchamber is maintained at a desired constant value and the centrate doesnot contact or overflow the seal.

In an alternate embodiment with the use of air line 143, the positivepressure level of the regulator acts on both the fluid in the reservoir122 and the area of the separation chamber above the centripetal pumpinlet. Because the positive pressure level of air applied in bothlocations is the same, the back pressure on the centrate discharge line(which is the pressure applied above the fluid in the reservoir) isvirtually always the same as the pressure in the air pocket at the topof the separation chamber. This enables the centripetal pump to operatewithout any net effect from either pressure.

In this exemplary embodiment the pump 120 and other system componentsare controlled responsive to the at least one control circuit 142 toassure that there is an adequate volume of air within the interior ofthe reservoir 122 at all times during centrate production. This assuresthat the reservoir provides the desired damping effect on changes incentrate discharge line pressure that might otherwise be caused by thepumping action of pump 120. This is done by maintaining the liquid inthe reservoir 122 at no higher than the upper liquid level detected bysensor 134. Further, the liquid level in the reservoir is controlled tobe maintained above the lower liquid level as sensed by sensor 136. Thisassures that the centripetal pump is not pumping air and aerating thecentrate.

In the exemplary arrangement the centrate flow out of the separationchamber is controlled through operation of the at least one controlcircuit. The exemplary control circuitry may operate the system duringprocessing conditions to maintain the incoming flow of cell suspensionby pump 108 to the separation chamber 90 at a generally constant rate,while the separation process is occurring with the motor 86 operating tomaintain the constant bowl speed to achieve the separation of thecentrate and the cell concentrate. The exemplary arrangement furtheroperates to maintain an ideally constant back pressure on the centratedischarge line from the centripetal pump while maintaining air in theseparation chamber to isolate the at least one seal 106 from thematerial being processed.

In the exemplary arrangement, the pressure maintained through operationof the regulator in the pressure damping reservoir is set atapproximately 2 kpa (0.29 psi) above atmospheric. In the exemplarysystem this pressure has been found to be suitable to assure that theseal integrity and isolation is maintained during all stages of cellsuspension processing. Of course it should be understood that this valueis exemplary and in other arrangements, other pressure values andpressure damping reservoir configurations, sensors and other featuresmay be utilized.

FIG. 20 shows schematically exemplary logic executed through operationof the at least one control circuit 142 in connection with maintainingthe desired pressure level in the centrate discharge tube and within thetop portion of the separation chamber. It should be understood that thecontrol circuits in some exemplary embodiments may perform numerousadditional or different functions other than those represented. Thesefunctions may include the overall control of the different processes andsteps for operation of the centrifuge in addition to the describedpressure control function. As represented in FIG. 20, in an initialsubroutine step 148, the at least one control circuit 142 is operativeto make a determination on whether the centrifuge operation is currentlyin a mode where centrate is being discharged from the separationchamber. If so, the at least one control circuit is operative to causethe centrate discharge pump 120 to operate to discharge centratedelivered through the centrate discharge line 118. This may be done bycausing operation of a motor of the pump. In the exemplary arrangement,the flow rate of the pump 120 may be a set value initially oralternatively may be varied depending on particular operating conditionsthat are determined through control circuit operation during theprocess. The operation of the centrate discharge pump is represented bya step 150.

The at least one control circuit is then operative to determine in astep 152 whether liquid is sensed at the high level of the high liquidlevel sensor 138. If so, this represents an undesirable condition. Ifliquid is sensed at the level of the sensor 138, the control circuitthen operates to take steps to address the condition. This may includeoperating the pump 120 to increase its flow rate and making subsequentdeterminations if the level drops within a period of time while thecentrifuge continues to operate. Alternatively or in addition, the atleast one control circuit may decrease the speed of pump 108 to reducethe flow of incoming material. If such action does not cause the levelto drop within a set period of time, additional steps are taken. Suchsteps may also include slowing or stopping rotation of the bowl 182.Such actions may also include stopping the operation of pump 108 so asto avoid the introduction of more suspension material into theseparation chamber. These steps which are generally referred to shuttingdown normal operation of the system are represented by a step 154.

If liquid is not sensed at the level of the high level sensor 138, theat least one control circuit is next operative to determine if liquid issensed at the upper liquid level of sensor 134. This is represented bystep 156. If liquid is sensed at the upper liquid level sensor, the atleast one circuit operates responsive to its stored instructions toincrease the speed and therefore the flow rate of discharge pump 120.This is done in an exemplary embodiment by increasing the speed of themotor that is a part of the pump. This is represented by a step 158.Increasing the flow rate of the pump causes the liquid level 147 in thepressure damping reservoir to begin to drop as more liquid is moved bythe pump 120.

If in step 156 liquid is not sensed at the upper liquid level of sensor134, the at least one control circuit then operates to make adetermination as to whether liquid is not sensed at the lower liquidlevel of sensor 136. This is represented by step 160. If the liquidlevel is not at the level of the sensor 136, the control circuitryoperates in accordance with its programming to control the pump 120 todecrease its flow rate. This is done in an exemplary embodiment byslowing the speed of the motor. This is represented by a step 162. Inthe exemplary arrangement, slowing the flow rate of the pump 120 causesthe liquid level 147 to begin rising in the pressure damping reservoir.In some exemplary arrangements if the level does not rise within thereservoir within a given time, the control circuitry may operate inaccordance with its programming to cause additional actions, such asactions associated with shut down step 154 previously discussed. Thecontrol circuitry of exemplary embodiments may operate to change thepumping rate of pump 120 to maintain the level 147 within the pressuredamping reservoir at a generally constant level between the levels ofsensors 134 and 136 during centrate production.

In the exemplary arrangement, maintaining the generally constantelevated pressure of sterile air over the liquid in the pressure dampingreservoir helps to assure that a similar elevated pressure isconsistently maintained in the centrate outlet line and at the sealwithin the separation chamber. Further in the exemplary arrangements,the pressure is enabled to be controlled at the desired level duringdifferent operating conditions of the centrifuge during which the bowlrotates at different speeds. This includes, for example, conditionsduring which the separation chamber is initially filled at a relativelyhigh rate through the introduction of cell suspension and during whichthe centrifuge rotates at a relatively lower speed. Pressure can also bemaintained during the subsequent condition of final fill in which theflow rate of cell suspension into the separation chamber occurs at aslower rate and during which the rotational speed of the bowl isincreased to a higher rotational speed. Further, positive pressure ismaintained as previously discussed during the feeding of the suspensioninto the bowl and during discharge of the centrate from the separationchamber. Further in exemplary embodiments, the at least one controlcircuit may operate to also maintain the positive pressure during thetime period that the concentrate is removed by having it pumped out ofthe separation chamber. Maintaining positive pressure within theseparation chamber during all of these conditions reduces the risk ofcontamination and other undesirable conditions which otherwise mightarise due to negative pressure (below atmospheric pressure) conditions.

Of course it should be understood that the features, components,structures and control methodologies are exemplary, and in otherarrangements other approaches may be used. Further, although theexemplary arrangement includes a system which operates in a batch moderather than a mode in which both centrate and concentrate arecontinuously processed, the principles hereof may also be applied tosuch other types of systems.

While the pressure damping reservoir is useful in exemplary embodimentsto help assure that a desired pressure level is maintained in the outlettube and the separation chamber, other approaches may also be utilizedin other exemplary embodiments. For example, in some arrangementspressure may be directly sensed and/or applied in the outlet tube, theseparation chamber or in other locations which correspond to thepressure in the separation chamber. In some arrangements, the flow rateof the discharge pump may be controlled so as to maintain the suitablepressure level. In still other arrangements, exemplary control circuitsmay be operative to control both the discharge pump and a pump thatfeeds suspension into the core and/or suitable valving or other flowcontrol devices so as to maintain suitable pressure levels. Suchalternative approaches may be desirable depending on the particularcentrifuge device being utilized and the type of material beingprocessed.

Thus the new centrifuge system and method of the exemplary embodimentsachieves the above stated objectives, eliminates difficultiesencountered in the use of prior devices and systems, solves problems andattains the desirable results described herein.

In the foregoing description certain terms have been used for brevity,clarity and understanding, however, no unnecessary limitations are to beimplied there from because such terms are for descriptive purposes andare intended to be broadly construed. Moreover, the descriptions andillustrations herein are by way of examples and the invention is notlimited to the exact details shown and described.

In the following claims any feature described as a means for performinga function shall be construed as encompassing any means capable ofperforming the recited function, and shall not be limited to thestructures shown herein or mere equivalents thereof.

Having described the features, discoveries and principles of the new anduseful features, the manner in which they are constructed, utilized andoperated, and the advantages and useful results attained, the new anduseful structures, devices, elements, arrangements, parts, combinations,systems, equipment, operations and relationships are set forth in theappended claims.

1. Apparatus comprising: a centripetal pump configured for use in acentrifuge operative to separate cell suspension into cell centrate andconcentrate, wherein the centrifuge includes a rotatable fluidcontaining core, wherein the core is rotatable about an axis in arotational direction, the centripetal pump comprising: a disk-shapedbody, wherein the disk-shaped body is concentric with the axis, whereinthe disk-shaped body includes a first plate and a second plate, anannular periphery, wherein the annular periphery is exposed to one ofeither centrate or concentrate within the core, and wherein the annularperiphery includes a an annular wall with an inlet opening, an annularoutlet opening, wherein the outlet opening is coaxial with the axis, anannular collection chamber, wherein the annular collection chamberextends radially outward of the outlet opening and verticallyintermediate of the first plate and the second plate, a curved volutepassage, wherein the curved volute passage extends within the body andradially between the inlet opening and the annular collection chamber,wherein the volute passage is bounded horizontally by opposed verticallyextending bounding walls, wherein one bounding wall extends transverselyof the rotational direction at the inlet opening and faces toward therotational direction, and wherein the one bounding wall is curvedcontinuously toward the rotational direction within the body radiallybetween the annular collection chamber and the inlet opening.
 2. Theapparatus according to claim 1 wherein both bounding walls are curvedtoward the rotational direction.
 3. The apparatus according to claim 1wherein the volute passage has a cross-sectional area, wherein thecross-sectional area is generally constant from the inlet to thecollection chamber.
 4. The apparatus according to claim 1 wherein thebody includes a pair of generally opposed volute passages.
 5. Theapparatus according to claim 4 wherein the volute passages are boundedvertically by respective upper and lower walls, wherein the first plateincludes the upper walls and the second plate includes the lower walls.6. The apparatus according to claim 5 wherein the first plate and thesecond plate are held in releasible engagement by at least one fastener.7. The apparatus according to claim 5 wherein one of the first plate andthe second plate includes recesses that bound three sides of eachpassage and two sides of the annular collection chamber, and wherein theother of the first plate and the second plate bounds one side of eachpassage and one side of the annular collection chamber.
 8. The apparatusaccording to claim 7 wherein the other of the first plate and the secondplate includes a planar circular face, and wherein the planar circularface vertically bounds the one side of each of the volute passages andthe one side of the annular collection chamber.
 9. The apparatusaccording to claim 7 wherein the inlets of the volute passages of thecentripetal pump are exposed to liquid cell centrate, and wherein thepump is operative to pump cell centrate from the core.
 10. The apparatusaccording to claim 7 wherein the inlets of the volute passages areexposed to cell concentrate in the core, and wherein the pump isoperative to pump cell concentrate from the core.
 11. The apparatusaccording to claim 1 and further comprising: the core, a furthercentripetal pump, wherein the further centripetal pump includes afurther disk-shaped body positioned coaxial with the axis, wherein thefurther disk-shaped body includes a further first plate and a furthersecond plate, and a further annular wall including a further annularperiphery, wherein the further annular wall includes a further inletopening, a further annular outlet opening, a further annular collectionchamber that extends radially outward from the further annular outletopening and vertically between the further first plate and the furthersecond plate, a further curved volute passage, wherein the furthercurved volute passage extends radially within the further body betweenthe further inlet opening and the further annular collection chamber,and wherein the further volute passage includes one further boundingwall, wherein the further bounding wall extends transversely of therotational direction at the further inlet opening and faces toward therotational direction, and wherein the one further bounding wall iscurved continuously toward the rotational direction within the furtherbody between the further annular collection chamber and the furtherinlet opening.
 12. The apparatus according to claim 11 wherein theannular periphery of the centripetal pump has a first diameter, andwherein the further centripetal pump has an annular periphery of asecond diameter, and wherein the second diameter is greater than thefirst diameter.
 13. The apparatus according to claim 12 wherein thefirst diameter and the second diameter have relative sizes that areoperative to avoid waves in centrate adjacent the inlet opening of thecentripetal pump.
 14. The apparatus according to claim 12 wherein thefurther centripetal pump is disposed in the core vertically above thecentripetal pump.
 15. (canceled)
 16. The apparatus according to claim 1and further comprising the core including the cell suspension, whereinthe inlet of the volute passage of the centripetal pump is exposed tocentrate, wherein the pump is operative to pump centrate from the core.17. The apparatus according to claim 1 and further comprising the coreincluding the cell suspension, wherein the inlet of the volute passageis exposed to concentrate in the core, wherein the pump is operative topump concentrate from the core.
 18. Apparatus comprising: a centripetalpump configured for use in a centrifuge operative to separate cellsuspension into cell centrate and concentrate, wherein the centrifugeincludes a rotatable fluid containing core, wherein the core isrotatable about a vertical axis in a rotational direction, thecentripetal pump comprising: a disk-shaped body, wherein the disk-shapedbody is concentric with the axis, wherein the disk-shaped body includesan annular periphery, wherein the annular periphery is exposed to one ofeither centrate or concentrate within the core, and wherein the annularperiphery includes a vertically extending peripheral wall with an inletopening, an annular outlet opening, wherein the outlet opening iscoaxial with the axis, an annular collection chamber, wherein theannular collection chamber extends radially outward from the annularoutlet opening and within the disk-shaped body, a curved volute passage,wherein the curved volute passage extends within the body and radiallyintermediate of the inlet opening and the annular collection chamber,wherein the volute passage is bounded horizontally by a verticallyextending bounding wall, wherein the vertically extending bounding wallextends transversely of the peripheral wall at the inlet opening and infacing relation of the rotational direction, and wherein the verticallyextending bounding wall is curved toward the rotational direction withinthe disk-shaped body.
 19. The apparatus according to claim 18 whereinthe volute passage is bounded by a pair of vertically extending boundingwalls, wherein both bounding walls are continuously curved toward therotational direction radially between the annular collection chamber andthe inlet opening.
 20. The apparatus according to claim 18 wherein thebody includes a plurality of angularly disposed curved volute passages.21. Apparatus comprising: a centripetal pump configured for use in acentrifuge operative to separate cell suspension into cell centrate andconcentrate, wherein the centrifuge includes a rotatable fluidcontaining core, wherein the core is rotatable about an axis in arotational direction, the centripetal pump comprising: a disk-shapedbody, wherein the disk-shaped body is concentric with the axis, whereinthe disk-shaped body includes an annular periphery, wherein the annularperiphery is exposed to centrate within the core, and wherein theannular periphery includes an inlet opening, an annular outlet opening,wherein the annular outlet opening is coaxial with the axis, an annularcollection chamber, wherein the annular collection chamber extendswithin the body and radially outward of the annular outlet opening, apassage, wherein the passage extends within the body and radiallybetween the inlet opening and the annular collection chamber, whereinthe centripetal pump is operative to pump centrate from the core, afurther centripetal pump axially disposed from the centripetal pumpwithin the core, the further centripetal pump comprising: a furtherdisk-shaped body, wherein the further disk-shaped body is concentricwith the axis, wherein the further disk-shaped body includes a furtherannular periphery, wherein the further annular periphery is exposed toconcentrate in the core, and wherein the further annular peripheryincludes a further inlet opening, a further annular outlet opening,wherein the further annular outlet opening is coaxial with the axis, afurther annular collection chamber, wherein the further annularcollection chamber extends within the further body radially outward ofthe further annular outlet opening, a further passage, wherein thefurther passage extends within the further body and radially between thefurther inlet opening and the further annular collection chamber,wherein the further centripetal pump is operative to pump concentratefrom the core.
 22. Apparatus comprising: a centripetal pump in acentrifuge that separates cell suspension into cell centrate and cellconcentrate, wherein the centrifuge includes a rotatable suspensioncontaining core, wherein the core is rotatable about an axis in arotational direction, the centripetal pump comprising: a disk-shapedbody, wherein the disk-shaped body is concentric with the axis, whereinthe disk-shaped body includes an annular periphery, wherein the annularperiphery is exposed to one of either the cell centrate or the cellconcentrate within the core, and wherein the annular periphery includesa wall with an inlet opening into which one of either the cell centrateor concentrate flows, an annular outlet opening, wherein the annularoutlet opening is coaxial with the axis, an annular collection chamber,wherein the annular collection chamber extends horizontally within thedisk-shaped body, fluidly between and radially outward of the annularoutlet opening, a curved volute passage, wherein the curved volutepassage extends radially within the body and fluidly between the inletopening and the annular collection chamber, wherein the volute passageis bounded horizontally by opposed vertically extending bounding walls,and wherein one bounding wall faces the rotational direction at theinlet opening and is curved within the body to extend further toward therotational direction from the annular collection chamber with increasedradial distance from the axis.
 23. The apparatus according to claim 22wherein the volute passage has a cross-sectional area, wherein thecross-sectional area is generally constant within the body from theinlet to the annular collection chamber.
 24. The apparatus according toclaim 22 and further including the core housing the cell suspension,wherein the inlet of the volute passage is exposed to the cellconcentrate in the core, and wherein the centripetal pump is operativeto pump the cell concentrate from the core.
 25. The apparatus accordingto claim 21 wherein the annular periphery of the disk-shaped bodyincludes a solid annular wall that extends parallel to the axis, whereina pair of angularly separated inlet openings extend through the solidannular wall, wherein the disk-shaped body further includes a pair ofcurved generally opposed volute passages, wherein each curved volutepassage extends within the body and radially between a respective inletopening and the annular collection chamber, wherein each volute passageis bounded horizontally by opposed vertically extending bounding walls,wherein each bounding wall is curved toward the rotational direction,wherein the further annular periphery of the further disk-shaped bodyincludes a further solid annular wall that extends parallel to the axis,wherein a pair of angularly disposed further inlet openings extendthrough the further solid annular wall, wherein the further disk-shapedbody further includes a pair of further curved volute passages, whereineach further curved volute passage extends within the further bodybetween a respective further inlet opening and the further annularcollection chamber, wherein each further volute passage is boundedhorizontally by a pair of opposed further vertically extending boundingwalls, wherein each further bounding wall is curved toward therotational direction.
 26. The apparatus according to claim 25 andfurther comprising the core and the cell suspension wherein the annularperiphery of the centripetal pump has a first diameter, and wherein theannular periphery of the further centripetal pump has a second diameter,wherein the first and second diameter have relative sizes that areoperative to avoid waves in centrate adjacent the inlet openings of thecentripetal pump.
 27. The apparatus according to claim 26 wherein thefurther centripetal pump is disposed in the core vertically above thecentripetal pump, wherein the further centripetal pump is operative topump cell concentrate from the core and the centripetal pump isoperative to pump cell centrate from the core.
 28. The apparatusaccording to claim 21 wherein the disk-shaped body comprises a firstplate and a second plate, wherein the annular periphery of thedisk-shaped body includes a solid annular peripheral wall that extendsparallel to the axis, wherein the peripheral wall includes an inletopening, a curved volute passage, wherein the curved volute passageextends within the body and radially intermediate of the inlet openingand the annular collection chamber, wherein the volute passage isbounded horizontally by a vertically extending bounding wall, whereinthe vertically extending bounding wall extends transversely of theperipheral wall at the inlet opening and in facing relation of therotational direction, and wherein the vertically extending bounding wallis curved toward the rotational direction within the disk-shaped body.